The research program will investigate nutritional and genetic risk factors for orofacial clefts. The specific aims for the 3-year study are to assess the potential "gene-environmental" interplay between genetic variation of 3 potential folate pathway genes (folate receptor gene, reduced folate carrier, and the N-acetal transferace 1 gene) among probands, maternal folic acid/multivitamin intake, and the risk of orofacial clefts. We propose to investigate the hypothesis that one or more of the 3 folate pathway genes are responsible for inadequate transport, accumulation, or metabolism of folate during critical stages of craniofacial development, making embryos susceptible to orofacial clefts even in the presence of clinically adequate maternal folate intake. By combining state-of-the-art molecular biology approaches, new genetic findings, and one of the largest case-control studies done on orofacial clefts, we will determine if maternal supplemental folic acid intake overcomes folate transport or metabolic dysfunction that may occur as a result of the embryo's genotypic variation for the 3 folate pathway genes and thus reduce the risk for orofacial clefting. The project has 3 collaborating research centers: California Birth Defects Monitoring Program, University of Nebraska, and Children's Hospital, Oakland. The research design will be case-control, including approximately 1200 cafes and controls, and will utilize maternal interview data in conjunction with genotyping of the folate receptor gene. Infants' DNA for genotyping will be obtained from residual newborn screening bloodspots, of which about 1250 DNA samples will be available for this study. Information on a variety of relevant covariates, such as parental cigarette smoking and the infant's genotype for transforming growth factor-alpha polymorphisms, will be available to analytically assess their contribution to risk for orofacial clefts. As one of the first attempts at investigating environmental and molecular genetic interactions in the epidemiology of congenital anomalies, this study endeavors to enhance our general understanding of the causes of orofacial clefts as well as our specific understanding of the apparent protective effect of folate supplementation on the occurrence of clefts. We observed a 50% reduction in risk for orofacial clefts among pregnant women who used vitamins. If this association ultimately proves causal, many of these anomalies will be preventable every year in the United States once it is understood what vitamin/diet component is important in facilitating the reduction in risk.